Fish2degCBL, Fish2CBR Primer (cytb-Fish), amplify 80 pb, Thomsen et al. 2012

PCR #7, Pag 130, Dec 04 2019

PCR reaction

• ~ 40 ng/ul DNA
• 0.5 ul primer (10um)
• 5 ul Hoststart Qiagen
• 12.5 ul total reaction
• 0.5 ul BSA

Cycling conditions (Thomsen et al. 2012)

• 7 min to 95°C
• 50 cycles: 94°C 30 s, 50°C 30 s, 72°C 20 s,
• 5 min to 72°C
• 4°C forever

Fragment sizes were verified on 1% agarose gels stained with GelRed 4ul PCR product 4ul 1kb 100 v, 2 hrs.

I reduced BSA volume to 0.5ul instead of 0.75ul. The amplification improved. Amplified one negative sample. So, I will not use the samples for the sequencing test.

Fish2deg, PCR